TYPING 6 MAJOR HEPATITIS-C VIRUS GENOTYPES BY POLYMERASE CHAIN-REACTION USING PRIMERS DERIVED FROM NUCLEOTIDE-SEQUENCES OF THE NS5 REGION

We recently developed a polymerase chain reaction (PCR) based method f or determining hepatitis C virus (HCV) genotypes using primers derived from nucleotide sequences of the NS5 region. Using this method, we de termined 5 hepatitis C virus (HCV) genotypes (1a, 1b, 2a, 2b and 3b) d etected in Japanese patients. Following the recent identification of g enotype 3a HCV in a patient from Pakistan currently living in Japan, w e improved the typing method used for detecting all of these 6 genotyp es. Eighteen of 1100 HCV RNA positive patients who tested negative for genotype by the original method also tested negative by the new metho d. Type 3a genotype HCV was thus shown to be very rare among Japanese patients with HCV infection. Nucleotide sequence and quantitative anal ysis of these negative samples revealed that negative results were due to mutations present on the nucleotide sequence where the 3' end of t he primers was situated due to a very low titer (less than 10(3)) of t he virus. The new single step genotyping method may be clinically usef ul as it allows a fast determination of all 6 genotypes present in Jap an.